A novel plasmid, pBSR2, was constructed by incorporating a strong lipase promoter and a terminator into the original pBD64. A mature lipase gene from Bacillus subtilis strain IFFI10210, an existing strain for lipase expression, was cloned into the plasmid pBSR2 and transformed into B. subtilis A.S.1.1655. Thus, an overexpression strain, BSL2, was obtained. The yield of lipase is about 8.6mg protein/g of wet weight of cell mass and 100-fold higher than that in B. subtilis strain IFFI10210. The recombinant lipase was puriWed in a three-step procedure involving ammonium sulfate fractionation, ion exchange, and gel Wltration chromatography. Characterizations of the puriWed enzyme revealed a molecular mass of 24 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, maximum activity at 43℃ and pH 8.5 for hydrolysis of p-nitrophenyl caprylate. The values of Km and Vm were found to be 0.37mM and 303 μmolmg-1·min-1, respectively. The substrate speciWcity study showed that p-nitrophenyl caprylate is a preference of the enzyme. The metal ions Ca2+, K+, and Mg2+ can activate the lipase, whereas Fe2+, Cu2+, and Co2+ inhibited it. The activity of the lipase can be increased about 48% by sodium taurocholate at the concentration of 7mM and inhibited at concentrations over 10mM.